Cells that retain their capacity to contribute to the germline of vertebrates can be used to make transgenic animals, to cryopreserve the genome and to make germline chimeras. In mice, embryonic stem cells have been used to make transgenic animals and to cryopreserve specific strains that are of interest to the biomedical research community. In chickens, primordial germ cells (PGCs) have been cultured for extended periods of time, their genome has been modified and they have been used to make germline chimeras.
In vertebrates, the germline is separated from the somatic tissues at an early stage of development. In birds, the germline may be separated after only a few cell divisions whereas in mice, PGCs are derived from the proximal epiblast cells around 7.2 dpc. During the course of gastrulation the murine PGCs move through the posterior primitive streak into the extra-embryonic region. Around 8.5 dpc the PGCs will migrate back into the embryo proper following a route through the gut mesentery and become incorporated into the somatic component of the developing gonad. In males, PGCs are incorporated into the developing seminiferous tubules and in females, they are incorporated into the ovarian cortex.
In birds, PGCs descend through the epiblast and are swept into the anterior extra-embryonic region known as the germinal crescent. This process is underway when the egg is laid (at approximately Stage XII using the staging system of Eyal-Giladi and Kochav Developmental Biology (1976) 49, 321-337)) and is completed by 18 hours of development (about Stage 4 to 5 using the staging system of Hamburger and Hamilton (Journal of Morphology (1955) 88, 49-92). From the germinal crescent, PGCs migrate into the nascent vascular system and travel to the gonadal ridge where they exit into the developing testis or ovary.
The migratory property which defines PGCs ends when they take up residence in the undifferentiated gonad. At this time, they are referred to as gonocytes to reflect their status as the primary germ cells in the gonadal environment. Gonocytes have a distinct gene expression profile that is reflected in their unique response to various effectors.
Certain aspects of this disclosure relate to a culture method for producing avian gonocytes, avian gonocytes produced by the method and methods for using the same.